This is a revised application. All changes are indicated by, a red line in the right margin. At the molecular level, acute myeloid leukemia (AML) is a heterogeneous disease. Recent advances with molecular-based risk stratification of AML and molecular-based therapeutics strongly suggest that elucidation of molecular mechanisms underlying each case of AML will have the greatest impact on increasing the cure rate of this disease. In the majority of AML cases, cytogenetics are either normal or only contain changes in chromosome number that limit one's ability to find leukemogenic gene fusions. Several years ago, our laboratory collaborated to discover a novel molecular defect found in 5-10 percent of AML cases with normal cytogenetics and in the majority of AML cases with trisomy 11 as a sole abnormality. The defect involves a partial tandem duplication (PTD) of the MLL gene, whereby exons 2-6 or 2-8 duplicate in tandem creating a unique self-fusions. Our laboratory has since performed an extensive characterization of the MLL PTD. We hypothesize that the MLL PTD represents a primary molecular defect in myeloid hematopoietic progenitor cells that is responsible, at least in part, for their leukemic transformation. We propose a series of in vitro and in vivo model systems to test this hypothesis and to define the genetic differences that specifically result from the MLL PTD. We have developed a targeting construct to create embryonic stem cells expressing the MLL PTD for in vitro differentiation studies as well as for creation of chimeric and heterozygous mice expressing the MLL PTD. Finally, we have utilized two techniques to study genome-wide genetic and epigenetic changes in leukemic tissue to better understand downstream effector molecules during malignant cell growth, and propose to use these technologies to better understand pathways critical to leukemogenesis in cells harboring the MLL PTD. Ultimately, we believe insights gained by the experiments proposed in this application will further our understanding of leukemogenesis and open up new therapeutic options for this subset of AML patients with a poor prognosis.